The dynein adaptor Lis1 plays a pivotal role in the oncogenesis of several cancers, including leukemia. In leukemia, Lis1 controls differential positioning of the mitotic spindle within the dividing cancer cell, resulting in the asymmetric cel divisions that allow these cells to divide away from their substratum and turn cancerous. Mutations in human Lis1 can also cause malformation of brain tissue, resulting in the devastating and fatal brain disease, Lissencephaly. The long-term interest of the Miller lab is to understand the microtubule-based mechanisms involved in orientation of the mitotic spindle, a process that requires the dynein motor protein. The Miller lab has shown that SUMO is important for positioning the mitotic spindle. The Miller lab recently established that the dynein adaptor Pac1p, the yeast homologue of Lis1, is modified by both ubiquitin and SUMO. Preliminary data from the Miller lab shows that sumoylation regulates Lis1/Pac1p. However, the exact role that these modifications play in regulating Lis1/Pac1p and dynein is not known. The objective of this proposal is to identify the specific mechanisms by which sumoylation regulate Lis1/Pac1p and to determine the extent to which the SUMO modification of Lis1 is conserved in human cells. The rationale is that by achieving a fuller understanding of this key sumo regulation in yeast cells and establishing its translational impact in human cells, researchers can advance similar investigations into many other tissue- specific disease models. The central hypothesis of this proposal is that sumoylation regulates Lis1 to control the dynein motor. This hypothesis will be tested by pursuing two specific aims using the yeast model organism. 1. Identify novel signal transduction mechanisms that regulate dynein. 2. Determine the role that SUMO targeted ubiquitin ligases (STUbLs) play in the activity of Lis1. A third Aim will characterize the SUMO modification in Lis1 in human cells. These Aims will be accomplished using a combination of biochemical, cell biological and genetic approaches. This work is significant because it will contribute to a new understanding of key mechanisms regulating microtubule-associated proteins and a major motor protein, aiding the development of improved cancer interventions. It will also provide insight into regulation of the sumoylated MAP tau, a MAP implicated in neurodegenerative diseases such as Alzheimer's. This proposal is innovative because the approach is based on a new paradigm of sumoylation regulation of the microtubule dependent process of spindle-positioning, guided by preliminary data the Miller lab has generated. This work supports the NIH mission to pursue fundamental knowledge about the behavior of systems and the application of that knowledge to extend healthy life. This research project will allow students at all levels to learn research strategies and critical thinking skillsin hands-on research, inspiring them to pursue biomedical careers at the next level. This project strengthens the overall research environment at OSU by facilitating several intra- disciplinary collaborations.